Parental depressive symptoms, children’s emotional and behavioural problems, and parents’ expressed emotion - Critical and positive comments

This longitudinal study examined whether mothers' and fathers' depressive symptoms predict, independently and interactively, children's emotional and behavioural problems. It also examined bi-directional associations between parents' expressed emotion constituents (parents' child-directed positive and critical comments) and children's emotional and behavioural problems. At time 1, the sample consisted of 160 families in which 50 mothers and 40 fathers had depression according to the Structured Clinical Interview for DSM-IV. Children's mean age at Time 1 was 3.9 years (SD = 0.8). Families (n = 106) were followed up approximately 16 months later (Time 2). Expressed emotion constituents were assessed using the Preschool Five Minute Speech Sample. In total, 144 mothers and 158 fathers at Time 1 and 93 mothers and 105 fathers at Time 2 provided speech samples. Fathers' depressive symptoms were concurrently associated with more child emotional problems when mothers had higher levels of depressive symptoms. When controlling for important confounders (children's gender, baseline problems, mothers' depressive symptoms and parents' education and age), fathers' depressive symptoms independently predicted higher levels of emotional and behavioural problems in their children over time. There was limited evidence for a bi-directional relationship between fathers' positive comments and change in children's behavioural problems over time. Unexpectedly, there were no bi-directional associations between parents' critical comments and children's outcomes. We conclude that the study provides evidence to support a whole family approach to prevention and intervention strategies for children's mental health and parental depression

The impacts of water pricing and non-pricing policies on sustainable water resources management: a case of Changa Pani water supply scheme in Pakistan

The impacts of water pricing and non-pricing policies on sustainable water resources management: a case of Changa Pani water supply scheme in Pakista

Reversal of Ischemic Cardiomyopathy with Sca-1⁺ Stem Cells Modified with Multiple Growth Factors

<div><p>Background</p><p>We hypothesized that bone marrow derived Sca-1⁺ stem cells (BM Sca-1⁺) transduced with multiple therapeutic cytokines with diverse effects will induce faster angiomyogenic differentiation in the infarcted myocardium.</p><p>Methods and Results</p><p>BM Sca-1⁺ were purified from transgenic male mice expressing GFP. Plasmids encoding for select quartet of growth factors, i.e., human IGF-1, VEGF, SDF-1α and HGF were prepared and used for genetic modification of Sca-1⁺ cells (^GFSca-1⁺). Scramble transfected cells (^ScSca-1⁺) were used as a control. RT-PCR and western blotting showed significantly higher expression of the growth factors in ^GFSca-1⁺. Besides the quartet of the therapeutic growth factors, PCR based growth factor array showed upregulation of multiple angiogenic and prosurvival factors such as Ang-1, Ang-2, MMP9, Cx43, BMP2, BMP5, FGF2, and NGF in ^GFSca-1⁺ (<i>p<</i>0.01 <i>vs</i>^ScSca-1⁺). LDH and TUNEL assays showed enhanced survival of ^GFSca-1⁺ under lethal anoxia (<i>p<</i>0.01 <i>vs</i>^ScSca-1⁺). MTS assay showed significant increased cell proliferation in ^GFSca-1⁺ (<i>p<</i>0.05 <i>vs</i>^ScSca-1⁺). For in vivo study, female mice were grouped to receive the intramyocardial injection of 15 μl DMEM without cells (group-1) or containing 2.5×10⁵^ScSca-1⁺ (group-2) or ^GFSca-1⁺ (group-3) immediately after coronary artery ligation. As indicated by <i>Sry</i> gene, a higher survival of ^GFSca-1⁺ in group-3 on day4 (2.3 fold higher <i>vs</i> group-2) was observed with massive mobilization of stem and progenitor cells (cKit⁺, Mdr1⁺, Cxcr4⁺ cells). Heart tissue sections immunostained for actinin and Cx43 at 4 weeks post engraftment showed extensive myofiber formation and expression of gap junctions. Immunostaining for vWF showed increased blood vessel density in both peri-infarct and infarct regions in group-3. Infarct size was attenuated and the global heart function was improved in group-3 as compared to group-2.</p><p>Conclusions</p><p>Administration of BM Sca-1⁺ transduced with multiple genes is a novel approach to treat infarcted heart for its regeneration.</p></div

Cell migration and angiogenic effects of CMD <i>in vitro</i>.

<p>(A)Trans-well cell migration assay using HUVECs showed that the number of migrating cells per microscopic field (200x) in response to ^GF-Sca1CMD was significantly higher compared to ^Sc-Sca1CMD as a control (15±2.38 <i>vs</i> 8±2.35, <i>p</i><0.01). Nuclei were stained with DAPI (blue). (B) Tube formation assay on matrigel using HUVECs treated with conditioned medium (CMD) from ^GFSca-1⁺ cells (^GF-Sca1CMD) or ^ScSca-1⁺ cells (^Sc-Sca1CMD). ^GF-Sca1CMD showed significantly higher number of branch points per microscopic field (100x) as compared to ^Sc-Sca1CMD.</p

(A) Real-time PCR based growth factor array showing up-regulation of multiple angiogenic and pro-survival factors including connexin-43 (Cx43), angiopoietin-1 & 2 (Ang-1 & Ang-2), matrix metalloproteinase-9 (MMP9), Bone morphogenetic protein 2 & 5 (BMP2 & 5), fibroblast growth factor-2 (FGF-2), and Nerve growth factor (NGF) in pooled ^GFSca-1⁺ cells compared to control Sca-1⁺ cells.

<p>(B; B1–B2) LDH release assay showed significant reduction of cell death in ^GFSca-1⁺ as compared to ^ScSca-1⁺ after 8 hours cultured under lethal anoxia (B1) and co-cultured cardiomyocytes after 3 hours under lethal anoxia (B2). (C) MTS assay was used to determine proliferative activity in vitro. The proliferation rate of ^GFSca-1⁺ group was significantly higher as compared to ^ScSca-1⁺ (0.59±0.06 vs. 0.42±0.04 absorbance at 490 nm) (D) TUNEL staining confirmed that 8 hours lethal anoxia caused higher TUNEL positivity in ^ScSca-1⁺ as compared to ^GFSca-1⁺ cells.</p

Masson trichrome staining was performed to evaluate the infarction size.

<p>Significant attenuation of infarct size was observed in ^GFSca-1⁺ group as compared to other groups. Indices of the left ventricular contractile function including LVEF and LVFS were significantly preserved in ^GFSca-1⁺ group at 4 weeks after cell engraftment as compared to other groups(**<i>P</i><0.01).</p

Flow cytometry of the purified mouse BM Sca-1⁺ cells for surface markers analysis.

<p>(A) unlabeled Sca-1⁺ cells (control; A1), labeled cells showing 96.6% Sca-1⁺ (A2),1.7% c-kit⁺(A3), and 1.4% CD45⁺(A4) cell populations. (B) Immunostaining of cells for Sca-1 antigen (red = Sca-1; blue = DAPI; magnification = 400x). (C–E) Representative figures of <i>in vitro</i> characterization of BM Sca-1⁺ cells transfected for expression of the respective transgene. RT-PCR (C) and western blot (D) for HGF, IGF-1, SDF-1α, and VEGF transgene expression <i>in vitro</i>. In each case, non-transfected native Sca-1⁺ cells were used as a control. Densitometry showed significantly up-regulated expression of the respective growth factor gene and protein in the transfected Sca-1⁺ cells 48 hours after transfection. (E) Fluorescence images of the Sca-1⁺ cells immunostained for respective growth factor HGF (E1, red), IGF-1(E2, red), SDF-1α (E3, red), VEGF (E4, red). DAPI was used to visualize the nuclei (blue, magnification = 400x).</p

Microscopic images from recipient mouse hearts 4 weeks post-transplantation.

<p>Fluorescence immunostaining of the heart tissue revealed increased angiomyogenic differentiation of the transplanted Sca-1⁺ in the infarct and peri-infarct regions 4 weeks after transplantation (GFP (green 5A1,5B1), α-actinin (red; <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0093645#pone-0093645-g005" target="_blank">Figure 5A</a>2) and vWF (red; <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0093645#pone-0093645-g005" target="_blank">Figure 5B</a>2)). Arrows show the merged color of red and green indicating the colocalization of α-actinin and GFP (5A3) or vWF and GFP (5B3). The nuclei were stained with DAPI (blue) (C) Immunostaining of the heart tissue revealed Cx43 (red) expression between^GFSca-1⁺ and the host cardiomyocytes (indicated by yellow arrows). (D) Cx43 (red) expression between ^GFSca-1⁺ and the co-cultured cardiomyocytes at day 4 after co-culture. (C–D) ^GFSca-1⁺ stem cells were prelabeled with DAPI (blue), and the nuclei were double stained with SYTOX (green).</p

miR-150 targets Cxcr4 and regulates MNC migration.

<p>(A) A putative target site of miR-150 highly conserved in the <i>Cxcr4</i> mRNA 3′-UTR as predicted by computational analysis. (B) miR-150 expression was decreased in BM-MNCs after AMI as validated by real-time PCR. (C) CXCR4 protein expression in MNCs significantly increased by transfection with miR-150 inhibitor. (D) BM-MNCs isolated from mice with LAD ligation enhanced migration capacity of BM-MNCs in response to SDF-1α as evaluated by transwell migration system. Transfection of wild type MNCs with anti-miR-150 also increased the number of migrating cells.</p

AMI increases the numbers of BM-MNCs and CXCR4 positive MNCs.

<p>(A) MNCs were isolated at 1, 3 and 5 days after left anterior descending artery ligation (LAD) and analyzed for molecular characterization. (B) The number of PB-MNCs decreased at one day after LAD ligation and increased gradually up to 5 days (n≥10 in each groups). (C) The percentage of CXCR4 positive cells in MNCs increased in both PB and BM after LAD ligation as compared to control. (n = 8 in control group and n = 6 in AMI group). (D) Densitometric analysis of CXCR4 positive MNCs counted after LAD ligation.</p